A gene trap cassette inserted into intron 1 of the gene. The gene trap cassette includes a splice adaptor at the 5â end with neomycin selection and poly(A) termination signals followed by another ES cell-only promotor that expresses the first exon of the Brutonâs tyrosine kinase gene with multiple termination codons to prevent expression of downstream fusion transcripts. Immunoblot analysis confirmed the absence of protein expression in isolated mouse embryonic fibroblasts (MEFs) and tissues from homozygous mutant mice.